The long-term objective of this research is to understand molecular aspects of the pathogenic process in prion diseases. These transmissible neurodegenerative disorders include Creutzfeldt-Jakob disease in humans, bovine spongiform encephalopathy in cattle and chronic wasting disease in cervids. The most intriguing aspect of prion diseases is the nature of the infectious agent that is composed of a misfolded form of the prion protein, PrPSc. This misfolded conformer is believed to self-replicate by the mechanism involving binding to the normal prion protein, PrPC, and forcing its conformational conversion to the sheet- rich PrPSc state. However, the mechanism of the PrPC ? PrPSc conversion and, especially, the structural aspects of this process remain poorly understood. The lack of information in this regard represents a critical gap in prion research, hindering efforts to understand the molecular basis of TSE diseases as well as development of effective strategies for pharmacological intervention. The overall goal of this Project is to gain insight into the structural basis of prion protein conversion and, especially, the structure of PrPSc conformer. The first specific aim seeks to gain insight into the molecular and structural basis of seeded conversion of prion protein to the infectious form, with the working hypothesis that the key initial event in this reaction is a conformational transition within a relatively short segment of the protein, from which ?-structure propagates into the surrounding regions. The second aim takes advantage of the recent seminal finding that highly infectious prions can be generated in vitro from the recombinant PrP in the presence of additional cofactors. We will use a battery of biophysical techniques to characterize these infectious protein aggregates at all levels of structural organization. The third aim seeks to determine conformational correlates of transmissibility and phenotypic variability of human prion diseases. This Research Project is one of the components of a larger Program Project. It relies on services provided by the Animal and Neuropathology Cores and involves close scientific interaction with two other Research Projects of this Program.